A variety of proteins, including both cell-surface receptors and soluble ligands, require dimerization for proper biological activity. Although dimerization may readily occur in the native biological milieu, it is often problematic for recombinant forms of many proteins, including soluble forms of receptors that are embedded in the cell surface in their native state.
Cell-surface receptors that must dimerize in order to transduce a signal include members of the receptor tyrosine kinase family, the tumor necrosis factor (TNF) receptor family, and the class 1 and class 2 cytokine receptor families. Platelet derived growth factor (PDGF) receptors, for example, dimerize upon ligand binding, resulting in autophosphorylation of tyrosine residues and initiation of intracellular signal transduction. Other examples of receptors that dimerize upon ligand binding include growth hormone receptors, interleukin-2 (IL-2) receptors, IL-3 receptors, IL-5 receptors, IL-6 receptors, granulocyte-macrophage colony stimulating factor (GM-CSF) receptors, oncostatin M (OSM) receptors, leukemia inhibitory factor (LIF) receptors, and ciliary neurotrophic factor (CNTF) receptors. Dimerized receptors include both homodimers and heterodimers. Some receptor polypeptides function as subunits within a plurality of cytokine receptor dimers. See, for example, Cosman, Cytokine 5:95-106, 1993.
Naturally occurring soluble forms of many cell-surface receptors are known. These soluble receptors correspond to the ligand-binding domains of their cell-surface counterparts. Soluble cytokine receptors inhibit cytokine responses and act as transport proteins. See, for example, Aggarwal and Puri, “Common and Uncommon Features of Cytokines and Cytokine Receptors An Overview,” in Aggarwal and Puri, eds., Human Cytokines: Their Role in Disease and Therapy, Blackwell Science, 1995, 3-24. It has been found that dimerization of soluble receptor polypeptides through the use of fusion proteins may enhance the binding properties of these soluble receptors so that they become therapeutically useful antagonists of their cognate ligands. Typical of such dimeric fusions are immunoglobulin fusions. See, for example, Sledziewski et al., U.S. Pat. Nos. 5,155,027 and 5,567,584; Jacobs et al., U.S. Pat. No. 5,605,690; Wallner et al., U.S. Pat. No. 5,914,111; and Ashkenazi and Chamow, Curr. Opin. Immunol. 9:195-200, 1997.
To date, immunoglobulin fusion technology has not provided a commercially viable means to produce heterodimeric proteins. Using currently available technology, co-expression of two different fusion polypeptides in a recombinant cell generally results in a mixture of both homodimers and heterodimers. The costs associated with recovery and purification of heterodimers from the mixture has limited the commercial application of this technology. Thus, there remains a need in the art for an efficient method of producing soluble, dimeric proteins, including soluble receptor dimers and soluble heterodimers.